THE ROLES OF A LONG NON-CODING RNA GM15441 IN LIVER ENERGY METABOLISM

Abstract

The majority of the mammalian genome comprises non-coding regions, where numerous long non-coding RNAs (lncRNAs) are transcribed. Although it has been revealed that some lncRNAs serve as key regulators in essential biological processes, the majority of their functions remain unknown, notably in liver metabolism. Analysis of our recent genome-wide screen data reveals that Gm15441, a lncRNA antisense to a protein-coding gene thioredoxin-interacting protein (Txnip), is the most significantly upregulated lncRNA in the fasting mouse liver. Given that certain antisense lncRNAs have been reported to regulate the expression of their sense genes, the roles of Gm15441 in regulating Txnip expression were addressed in this work. Furthermore, the fact that Txnip is a prominent regulator of liver metabolism prompted us to investigate the functional role of the Gm15441-Txnip axis in liver metabolic homeostasis.We first investigated the response of Gm15441 and Txnip to in vivo metabolic challenges, such as fasting and refeeding, and in vitro metabolic stimuli, such as insulin and key metabolic transcription factors. Next, in mouse hepatocytes, we assessed the regulation of Txnip expression by Gm15441 and the underlying mechanism. Lastly, using an adenovirus-mediated liver-specific overexpression approach, we determined whether Gm15441 regulates Txnip expression in the mouse liver and modulates key aspects of liver metabolism, including glucose and lipid metabolism. We found that Gm15441 is most robustly upregulated in the mouse liver upon fasting. In addition, Gm15441 and Txnip expression levels were shown to respond similarly to various metabolic signals both in vivo and in vitro, although their functions were predicted to be opposed. Furthermore, knockdown of Gm15441 by siRNA showed a trend of increasing Txnip protein expression in hepatocytes, while overexpression of Gm15441 by adenoviral gene delivery dramatically reduced Txnip protein expression both in vitro and in vivo. Mechanistically, we showed that Gm15441 directly binds to Txnip at RNA levels and inhibits Txnip translation. Moreover, the 207-709 nucleotides of Gm15441, which is complementary to the Txnip 5’end, mediate the reduction of Txnip protein expression. Lastly, we found that liver-specific overexpression of Gm15441 by adenovirus regulates glucose and lipid metabolism by decreasing blood glucose and plasma triglyceride levels while increasing plasma ketone body levels. In conclusion, our study demonstrated that lncRNA Gm15441 is a potent translational inhibitor of Txnip and a critical regulator in the liver metabolic homeostasis. This study characterizes both gene modulation and biologic functional role of Gm15441 and reveals the therapeutic potential of lncRNA Gm15441 for the treatment of glucose and lipid metabolic disorders.